Evaluation of L-carnitine supplementation during equine oocyte in vitro maturation

Abstract

Efficiency of in vitro embryo production in the horse remains limited, in part due to poor quality of in vitro matured equine oocytes. Equine in vitro maturation (IVM) is suboptimal, with only ~60% of immature oocytes maturing in culture, and those that mature often exhibit reduced quality compared with oocytes matured in vivo. A suboptimal culture environment likely contributes to these outcomes. Commonly used maturation media fail to recapitulate the in vivo maturation environment, lacking several metabolites present in follicular fluid. One such metabolite is L-carnitine, an essential cofactor for FAO. With emerging evidence that equine oocytes may rely on lipid metabolism for energy during maturation, L-carnitine therefore represents a potential strategy to improve IVM outcomes. The objective of this study was to determine whether L-carnitine supplementation during equine IVM enhances oocyte maturation, improves indicators of oocyte quality, and alters metabolic activity of the cumulus-oocyte complex (COC). Immature oocytes were recovered by transvaginal ultrasound-guided follicle aspiration and randomly assigned to standard maturation media, 2mM L-carnitine, and 4mM L-carnitine supplemented treatments. Following culture, oocytes were evaluated for nuclear maturation and cumulus expansion. Mitochondrial membrane potential, mitochondrial distribution, and reactive oxygen species (ROS) were also assessed using fluorescence microscopy. Alterations in metabolic activity were evaluated by GC-MS analysis of spent culture media. L-carnitine did not significantly alter maturation rates; however, cumulus expansion was significantly enhanced in L-carnitine treated groups. ROS levels and mitochondrial distribution were not affected by treatment, while mitochondrial membrane potential was significantly reduced in the 4mM treatment as compared to standard maturation media. Analysis of spent media revealed increased lactate production in L-carnitine treated groups relative to fresh media, suggesting altered glycolytic activity during maturation. These findings represent the first evaluation of L-carnitine supplementation during equine IVM and suggest that L-carnitine may influence oocyte quality and COC metabolic activity during maturation, potentially improving in vitro embryo production outcomes.